Int.J.Curr.Microbiol.App.Sci (2015) Special Issue-1: 29-37
ISSN: 2319-7706 Special Issue-1 (2015) pp. 29-37
http://www.ijcmas.com
Original Research Article
Trends of Bacterial Meningitis and Sensitivity Profile of Isolates:
A Five Year Study
Asfia Sultan*, Abida Khatoon, Meher Rizvi, Fatima Khan, Asma Husein Roohani,
Huma Naim, Indu Shukla and Haris M Khan
Department of Microbiology, JNMCH, AMU, Aligarh, India
*Corresponding author
ABSTRACT
Keywords
Meningitis,
Antimicrobial
resistance,
MRSA,
ESBL,
HLAR
Acute bacterial meningitis (ABM) is a significant worldwide cause of death in
adults. It is important to know the regional bacterial etiology along with their
sensitivity profile to allow optimum management of patients. This study was
undertaken to evaluate the trends in etiology of bacterial meningitis and
antimicrobial resistance pattern of pathogens over a period of 5 years. The study
was performed from Oct 2009 to Oct 2014. CSF samples were collected from all
patients suspected of meningitis and cultured on chocolate agar, blood agar and
MacConkey agar. Antimicrobial susceptibility testing was done using Kirby Bauer
disc diffusion method. Detection of MRSA, HLAR, ESBL, AmpC and MBL was
done. Total 2,313 CSF samples were received, of these 262(11.32%) were culture
positive. Majority of the patients were from paediatric age group. Female to male
ratio was 1:1.2. Gram positive bacteria 152(58.0%) were the predominant pathogen
followed by 75(28.6%) Gram negative bacilli. The most common bacteria isolated
wasStaphylococcus aureus88 (33.6%) followed by Citrobacter 38(14.5%) and
32(12.2%) Acinetobacter species. During five year study period there is decrease in
meningitis cases [2010(27.9%), 2011(27.5%), 2012(21.8%), 2013(4.9%),
2014(10.7%)] and constant decrease in number of S. pneumonae. Vancomycin and
amikacin was the most sensitive antibiotic for gram positive bacteria. Gram
negative bacilli showed good sensitivity for amikacin. Methicillin resistance in S.
aureus was 48.4%, HLAR among the Enterococci was 70% while among the
Enterobacteriaceae ESBL and AmpC production was 21.67% and 52%
respectively. The high prevalence of drug resistant pathogens should be dealt with
by rational use of antimicrobials. Measures should be taken to control the spread of
these resistant strains before they reach the alarming levels.
Introduction
the mortality rate due to ABM remains
significantly high in India and other
developing countries, ranging from 16 32%
(Mani et al., 2007). Optimum management
Acute bacterial meningitis (ABM) remains a
major cause of mortality and long-term
neurological sequelae worldwide. Despite
the availability of potent newer antibiotics,
29
Int.J.Curr.Microbiol.App.Sci (2015) Special Issue-1: 29-37
of patients requires determination of
regional bacterial etiology along with their
respective sensitivity profiles. According to
reports released by the World Health
Organization
(WHO),
Neisseria
meningitides, Streptococcus pneumoniae
and Haemophilus influenzae Type b
represent the triad responsible for over 80%
of all cases of bacterial meningitis
worldwide (WHO, 2005).
After the naked eye examination for the
presence
of
turbidity,
microscopic
examination was done by Gram s staining of
the centrifuged deposit of CSF. Immediately
after centrifugation of CSF, culture was
done on a plate of chocolate agar, 5% sheep
blood agar, Mac Conkeyagarand a tube of
brain heart infusion broth. These plates were
incubated for 24-48 hours in humid air plus
5-10% CO2 at 37 C. Cultures showing
growth were identified byusing standard
biochemical test (Collee 2006b).
There have been several published studies
regarding meningitis conducted in hospitals
in the developed countries but there is
paucity of data collected from developing
countries,
particularly
the
Indian
subcontinent.
Regional
information
regarding trends in terms of etiology and
antimicrobial susceptibility are essential for
correct and timely management of
meningitis. Keeping the current dismal
scenario in mind, the following study was
undertaken to evaluate the changing trends
in etiology and antimicrobial resistance
pattern of pathogens over a period of five
years (2009 2014) in a tertiary care hospital
of north India with emphasis on the
prevalence
of
methicillin
resistant
Staphylococcus aureus (MRSA), high level
aminoglycoside resistance in Enterococcus
species (HLAR), extended spectrum
lactamases (ESBL), Amp C and metallobetalactamases (MBL).
Antimicrobial susceptibility testing
Antibiotic susceptibility testing was
performed by Kirby Bauer disc diffusion
method as per CLSI guidelines on Mueller
Hinton agar (CLSI 2008). Gram positive
isolates were tested against amikacin
(30µg), gentamicin (10µg), levofloxacin
(5µg), sparfloxacin (5µg), erythromycin
(15µg), vancomycin (30µg), oxacillin (1µg),
tobramycin (10µg), amoxycillin (30µg).
Gram negative isolates were tested against,
amikacin (30µg), gentamicin (10µg),
levofloxacin (5µg), sparfloxacin (5µg),
ceftriaxone (30µg), cefoperazone (75µg),
cefoperazone-sulbactam
(75µg,
1:1),
cefixime
(5µg)
ceftriaxone-salbactam
(30/15µg), piperacillin (100µg), piperacillintazobactum (100:10µg), cefotaxime (30µg)
and tobramycin (10µg), cefoxitin (30µg),
imipenem (10µg). All discs were obtained
from HiMedia, India.
Material and Methods
This retrospective study was performed in
the Department of Microbiology, JNMCH
between January 2010 and December 2014.
Cerebrospinal fluid (CSF) samples were
collected from all patients suspected for
meningitis. 3-5 ml of CSF was collected by
lumbar puncture taking all the aseptic
precautions (Collee, 2006a). The specimens
were processed immediately and in case of
delay, they were kept in the incubator at
37 C.
Detection of extended spectrum and
AmpC beta lactamases
Screening of possible ESBL production was
done by using ceftriaxone (30 g) and
cefoparazone (75 g). Isolates showing zone
diameter less than 25 mm for ceftriaxone
and less than 19 mm for cefoparazone were
subsequently confirmed by disc potentiation
30
Int.J.Curr.Microbiol.App.Sci (2015) Special Issue-1: 29-37
test using cefoparazone and cefoparazonesalbactam combination (Rizvi et al. 2009).
Organisms sensitive to cefoxitin and
resistant to cefoperazone-salbactam and
piperacillin-tazobactam combination were
considered to be Amp C producers (CLSI
2008).
meningitidis, S. pneumoniae (Schlech et al.,
1985)
and
Listeria
monocytogenes
(Schuchat et al., 1997), the relative
incidence of meningitis caused by these
agents is less in South East Asia (Bhat et al.,
1991).
During the 5 year study period, total 2,313
CSF samples were received from patients
suspected of meningitis. Out of these
274(11.8%) were culture positive. Culture
positive cases have increased (11.8%) as
compared to our previous reports (Khan et
al. 2015). Although there was a constant
decrease in culture positive meningitis cases
over the five year study period [2010:
76(27.7%), 2011: 68(24.8%), 2012:
61(22.3%), 2013: 36(13.1%), and 2014: 33
(12.0%)] (Graph 1). The female-to-male
ratio in our patients was 1:1.2. This indicates
that males are prone to have bacterial
meningitis. Similar gender discrepancy in
meningitis is found in other countries
(McCormick et al., 2013).
Detection of metallo-betalactamases
Imipenem resistant isolates were tested for
metallo-beta-lactamases (MBL) production
by modified Hodge test and Double Disc
synergy test using EDTA (Lee et al. 2007)
Screening for methicillin resistance using
oxacillin disc test
Test was performed on Muller Hilton agar
with 4% NaCl using Oxacillin 1µg disc.
Any decrease in sensitivity zone was
considered as resistant.
HLAR resistance
In case of Enterococcus, HLAR was
detected using High content gentamycin
(120µg) and streptomycin (300µg) (Murray
2003).
Majority of the patients were from paediatric
age group (<12years). Prevalence was
predominantly in the 1 5 year age group
with 77(28.1%) cases followed by 5 15year
53(19.3%) cases. Among the infant group,
majority of cases 46(16.8%) were between
0-1month of age, while between 1 and 3
month 28(10.2%) cases, 3 6month 21(7.7%)
cases, 6 12month 34(12.4%) cases were
positive. Among adults the distribution was
as follows: 35 45year 5(1.8%), 15 25 year
4(1.5%), 25 35year 4(1.5%) and >45year
2(0.7%). (Graph 2 describes aetiology with
age).
Result and Discussion
Acute bacterial meningitis continues to be a
formidable illness with high morbidity and
mortality among children and adults in India
which warrants early diagnosis and
aggressive therapy. The main pathogens of
bacterial meningitis are known to be
different in different age groups, eras, and
geographic
areas.
The
choice
of
antimicrobial therapy is based on the most
common pathogen prevalent in a particular
geographical area and age group and their
antibiotic susceptibility pattern. Though the
common pathogens associated with bacterial
meningitis in the west are H. influenzae, N.
Etiology
Gram
positive
bacteria
were
the
predominant isolates in majority of cases of
meningitis 164(59.9%) while 110(40.1%)
were gram negative bacilli. In our study S.
31
Int.J.Curr.Microbiol.App.Sci (2015) Special Issue-1: 29-37
aureus has emerged as the most common
pathogen causing acute bacterial meningitis
in all the five years accounting for a total of
almost 35.8% of all isolates followed by
Citrobacter species which were way behind
at 13.9% and Pseudomonas aeruginosa at
13.1%. Frequency of S. pneumoniae
isolation has increased 13(4.7%) as
compared to our previous reports (Khan et
al. 2011) but prevalence is low as reported
in another Indian study (Bareja et al. 2013).
There was constant decrease in the
prevalence of S. pneumoniae during the
consecutive years [2010, 10(0.36%), 2011,
2(0.73%), 2012, 1(0.36%), 2013, 0(0%),
2014, 0(0%)].
Klebsiella and S. aureus. CONS was seen
mostly in the 1 5years age groups. Most
common pathogen between 1-5 years of age
was Citrobacter species. S. pneumoniae
cases were clustered between 0-5 years age
group. All three cases of Listeria
monocytogenes occurred between 0 and 1
month of age. Among 5 15 years S. aureus
was the most common isolate.
The difference in etiology from the
temperate west may be due to the fact that
India is a semitropical country where hardy
bacteria like S. aureus, CONS and gram
negative bacilli flourish and the relatively
more fragile bacteria like H. influenzae, N.
meningitidis and S. agalactiae in
comparison do not have a survival
advantage (Khan et al., 2011). These results
highlight the very different etiological
profile in India in comparison to that of the
west.
Studies have shown a similar increase in the
incidence of staphylococcal infection and a
decrease in the incidence of Streptococcus
pneumoniae (Huang et al., 2005, Pederson
et al., 2006). H. influenzae, N. meningitidis
and S. agalactiae were not isolated in our
study. Other Indian studies have also quoted
a low isolation rate of these pathogens (Bhat
et al., 1991, Kabra et al., 1991). However, in
this study Listeria monocytogenes was
isolated in 3(1.1%) of cases and 1
Coryneform species was isolated.
An increasing drug resistance was seen
among the Staphylococcal species and the
Enterococcus while Streptococcus species
maintained a uniform sensitivity throughout
the study period. A constant decline was
noticed in isolates of S. aureus against
fluoroquinolones from 20(83.3%) in 2010 to
6(60%) in 2014. Prevalence of MRSA was
nearly 30 35% in the study period except in
2011 when 15.3% isolates were methicillin
resistant. Frequency of MRSA and HLAR
has decreased as compared to our previous
reports (Khan et al. 2015). However, this
incidence of MRSA and HLAR is high as
compared to study by Bareja et al. (2013).
Fortunately, no vancomycin resistance was
detected in S. aureus or Enterococcus
species. Certain studies have reported low
level resistance to vancomycin in
Staphylococcal isolates (Gad et al., 2010)
which indicate an upcoming resistance to
even this reserve drug. Around 80% 90%
isolates were sensitive to aminoglycosides.
The other important emerging pathogens
were CONS and Enterococcus species
accounting for 9.9% and 8.0% cases of
meningitis respectively. Recent reports from
India have also shown similar frequency of
CONS in bacterial meningitis cases (Bareja
et al., 2013). An increase in number of
CONS and Enterococcus species in the
earlier part of study pointed their recent
emergence but incidence was decreased in
later years.
Across all age groups S. aureus, Citrobacter
species, E. coli, P. aeruginosa, and E.
faecalis were most commonly isolated.
Citrobacter species has emerged as most
common pathogen in neonates replacing
32
15(78.9)
16(61.6)
11(57.8)
6(60)
8(80)
6(100)
8(80)
1(100)
10(100)
2(100)
1(100)
5(33.3)
0(0)
1(25)
-
22(84.6)
13(73.6)
4(40)
4(40)
4(66.6)
7(70)
1(100)
-
15(62.5)
13(54)
18(94.7)
Cephalos
porins
Erythrom
ycin
33
*High content gentamycin and streptomycin to detect HLAR
-
3(75)
3(100)
-
-
-
-
-
-
-
-
-
2(50)
2(66.6)
8(53.3)
-
-
-
1(100)
2(100)
10(100)
-
1(100)
-
2(50)*
1(33.3)*
7(46.6)*
-
-
-
1(100)
1(50)
8(80)
-
1(100)
10(100)
6(100)
8(80)
7(70)
17(89.4)
22(84.6)
19(100)
20(83.3)
Aminogly
cosides
2014
2013
2012
2011
2010
2014
2013
2012
2011
2010
2014
2013
2012
2011
2010
2014
2013
2012
2011
2010
Organism
S. pneumoniae
13(86.6)
-
-
-
1(100)
-
-
-
1(100)
8(80)
5(83.3)
8(80)
6(60)
20(76.9)
13(68.4)
16(84.7)
20(83.3)
olones
Flouroquin
CONS
2(100)
10(100)
-
1(100)
8(80)
5(83.3)
8(80)
6(60)
13(68.4)
17(65.3)
16(84.7)
16(66.6)
Oxacillin
S. aureus
10(100)
6(100)
10(100)
10(100)
19(100)
26(100)
19(100)
24(100)
Vancom
ycin
Int.J.Curr.Microbiol.App.Sci (2015) Special Issue-1: 29-37
Table.1 Antimicrobial susceptibility pattern among Gram Positive cocci
E. faecalis
0(0)
0(0)
4(57.1)
10(83.3)
3(100)
3(100)
4(57.1)
7(100)
3(100)
7(100)
12(100)
0(0)
3(100)
7(100)
-
-
5(55.5)
-
-
9(100)
1(100)
1(100)
1(9)
4(100)
4(100)
11(100)
4(50)
8(100)
6(75)
1(50)
2(100)
8 (100)
0(0)
1(100)
2(66.6)
4(66.7)
6(100)
34
3(42.9)
3(100)
5(71.4)
6(85.7)
3(100)
5(71.4)
7(58.3)
1(14.2)
0(0)
-
3(33.3)
3(42.8)
2(66.6)
6(85.7)
6(50)
3(42.8)
0(0)
3(42.8)
3(33.3)
8(72.7)
3(37.5)
1(33.3)
-
-
1(100)
3(75)
5(62.5)
1(50)
1(100)
4(66.6)
1(50)
Aminogly
cosides
2014
2013
2012
2011
2010
2014
2013
2012
2011
2010
2014
2013
2012
2011
2010
2014
2013
2012
2011
2010
Organism
Citrobacter
4(33.3)
2(28.5)
0(0)
0(0)
4(44.4)
5(45.5)
4(50)
1(33.3)
-
-
1(100)
0(0)
3(37.5)
0(0)
0(0)
3(50)
1(50)
Flouroquino
lones
Klebseilla
1(9)
3(37.5)
0(0)
-
-
1(100)
2(50)
3(37.5)
1(50)
0(0)
3(50)
0(0)
Cephalosporins
E. coli
3(100)
0(0)
Cephalosporins+
inhibitor combination
2(100)
Carbepenems
Int.J.Curr.Microbiol.App.Sci (2015) Special Issue-1: 29-37
Table.2 Trend of antimicrobial susceptibility among Gram negative isolates
Pseudomonas sp
Int.J.Curr.Microbiol.App.Sci (2015) Special Issue-1: 29-37
Graph.1 Year wise distribution of bacterial isolate
Graph.2 Age wise distribution of bacterial isolates
Thus aminoglycosides are still the most
effective group of antimicrobials barring
glycopeptides (vancomycin) against GPCs.
increased as compared to our previous report
(Khan et al. 2011). Another interesting
finding is the alarming rise in AmpC
producers among all GNRs which was not
there in our previous report (Khan et al.,
2011). In E. coli, resistance was maximum
in 2010 and 2012 when all the isolates were
AmpC producers. Strains of Proteus were
resistant to all the drugs tested except
carbepenems. Unlike GPCs aminoglycosides
resistance has emerged significantly among
GNRs.
Among GNRs Citrobacter species were the
most resistant pathogens with 44.4% ESBL
production. There was an increase in ESBL
from 37.5% in 2010 to 44.4% in 2014 and
AmpC producers from 25% in 2010 to
100% in 2014 respectively. Resistance to
aminoglycosides and fluoroquinolones was
almost constant with slight decrease (27.3%)
in 2011. Frequency of ESBL production has
35
Int.J.Curr.Microbiol.App.Sci (2015) Special Issue-1: 29-37
Amongst the nil-fermenters, poor sensitivity
was shown against cephalosporins (28.5%)
in 2010 which improved to 71.4% in 2012
whereas
cephalosporin+
inhibitor
combination showed sensitivity greater than
80% except during 2010 and 2014 where a
dip in the sensitivity was noted (57.1%). An
interesting finding was improvement in
fluoroquinolone sensitivity from 14.2% in
2010 to 85.7% in 2014. This could be due
to less use of fluoroquinolones in treatment
of meningititis in our setting. Acinetobacter
species were sensitive to all the antibiotics
tested. MBL production was not detected in
any of the strains isolated.
Bhat, B.V., Verma, I.C., Puri, R.K.,
Srinivasan, S., Nalini, P. 1991. A
profile of pyogenic meningitis in
children. J. Indian. Med. Assoc., 89:
224 227.
Clinical and Laboratory Standards Institute
2003. Performance standards for
antimicrobial susceptibility testing:
eighteenth informational supplement:
Approved
standards
M100-S18.
Clinical and Laboratory Standards
Institute, Baltimore, USA. 2008.
Collee, J.G., Fraser, A.G., Marmion, B.P.,
Simmons, A. 2006a. Laboratory
strategy in the diagnosis of infective
syndromes. In: Collee, J.G., Duguid,
F.P., Fraser, A.G., Marmion, B.P.,
Simmons, A. (Eds). Mackey and
McCartney
practical
medical
microbiology, 14th edn. Elsevier,
NewDelhi, India. Pp. 77.
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2006b. Simmons tests for the
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J.G., Duguid, F.P., Fraser, A.G.,
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149.
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Epidemiology
and
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from different infectious diseases.
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Huang, C.R., Lu, C.H., Wu, J.J. 2005.
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staphylococcal
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in
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The epidemiologic trends of ABM should be
examined frequently because any change
may influence the choice of initial empiric
antibiotic therapy greatly. These results
revealed the altered trend in etiology of
meningitis cases and also signify the
upcoming levels of drug resistance amongst
the gram positive and the gram negative
microbes. Stress should be given to control
the spread of these resistant strains before
they reach the alarming levels in this region
and on the restrained & rationale use of
antimicrobials both in and outside the
hospital.
Acknowledgment
This study was extracted from the thesis
written by Ms. Akramashrafizaveh which
was approved by Shiraz University of
Medical Sciences.
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